Paramagnetic probes in NMR and EPR studies of membrane enzymes

The applications of paramagnetic probes to problems of structure and mechanism are discussed from the point of view of the membrane enzymologist. Problems unique to membrane systems are discussed, and a variety of nuclear and paramagnetic probes are evaluated. Three membrane ATPase (kidney (Na+ + K+)-ATPase, Ca2+-ATPase from sarcoplasmic reticulum and Mg2+-ATPase from kidney) are used to describe the types of experiments which can be done, the information which can be obtained and the limitations involved. Nuclear relaxation studies employing 1H, 7Li+, 31P and 205Tl+ nuclei are described. The advantages and disadvantages of Mn2+, Gd3+ and Cr3+ as paramagnetic probes are discussed in terms of the three ATPases. The theory and interpretation of Mn2+ and Gd3+ EPR spectra are evaluated in studies with the (Na+ + K+)-ATPase and Ca2+-ATPase, respectively.     

Discriminant haplotypes of avirulence genes of Phytophthora sojae lead to a molecular assay to predict phenotypes

The soybean–Phytophthora sojae interaction operates on a gene-for-gene relationship, where the product of a resistance gene (Rps) in the host recognizes that of an avirulence gene (Avr) in the pathogen to generate an incompatible reaction. To exploit this form of resistance, one must match with precision the appropriate Rps gene with the corresponding Avr gene. Currently, this association is evaluated by phenotyping assays that are labour-intensive and often imprecise. To circumvent this limitation, we sought to develop a molecular assay that would reveal the avirulence allele of the seven main Avr genes (Avr1a, Avr1b, Avr1c, Avr1d, Avr1k, Avr3a, and Avr6) in order to diagnose with precision the pathotypes of P. sojae isolates. For this purpose, we analysed the genomic regions of these Avr genes in 31 recently sequenced isolates with different virulence profiles and identified discriminant mutations between avirulence and virulence alleles. Specific primers were designed to generate amplicons of a distinct size, and polymerase chain reaction conditions were optimized in a final assay of two parallel runs. When tested on the 31 isolates of known virulence, the assay accurately revealed all avirulence alleles. The test was further assessed and compared to a phenotyping assay on 25 isolates of unknown virulence. The two assays matched in 97% (170/175) of the interactions studied. Interestingly, the sole cases of discrepancy were obtained with Avr3a, which suggests a possible imperfect interaction with Rps3a. This molecular assay offers a powerful and reliable tool to exploit and study with greater precision soybean resistance against P. sojae.     

Receptor-like cytoplasmic kinases are pivotal components in pattern recognition receptor-mediated signaling in plant immunity

Innate immunity is generally initiated with recognition of conserved pathogen-associated molecular patterns (PAMPs). PAMPs are perceived by pattern recognition receptors (PRRs), leading to activation of a series of immune responses, including the expression of defense genes, ROS production and activation of MAP kinase. Recent progress has indicated that receptor-like cytoplasmic kinases (RLCKs) are directly activated by ligand- activated PRRs and initiate pattern -triggered immunity (PTI) in both Arabidopsis and rice. To suppress PTI, pathogens inhibit the RLCKs by many types of effectors, including AvrAC, AvrPphB and Xoo1488. In this review, we summarize recent advances in RLCK-mediated PTI in plants.     

Nod1-mediated lipolysis promotes diacylglycerol accumulation and successive inflammation via PKCδ-IRAK axis in adipocytes

Chronic inflammation contributes to obesity mediated metabolic disturbances, including insulin resistance. Obesity is associated with altered microbial load in metabolic tissues that can contribute to metabolic inflammation. Different bacterial components such as, LPS, peptidoglycans have been shown to underpin metabolic disturbances through interaction with host innate immune receptors. Activation of Nucleotide-binding oligomerization domain-containing protein 1 (Nod1) with specific peptidoglycan moieties promotes insulin resistance, inflammation and lipolysis in adipocytes. However, it was not clear how Nod1-mediated lipolysis and inflammation is linked. Here, we tested if Nod1-mediated lipolysis caused accumulation of lipid intermediates and promoted cell autonomous inflammation in adipocytes. We showed that Nod1-mediated lipolysis caused accumulation of diacylglycerol (DAG) and activation of PKCδ in 3T3-L1 adipocytes, which was prevented with a Nod1 inhibitor. Nod1-activated PKCδ caused downstream stimulation of IRAK1/4 and was associated with increased expression of proinflammatory cytokines such as, IL-1β, IL-18, IL-6, TNFα and MCP-1. Pharmacological inhibition or siRNA mediated knockdown of IRAK1/4 attenuated Nod1-mediated activation of NF-κB, JNK, and the expression of proinflammatory cytokines. These results reveal that Nod1-mediated lipolysis promoted accumulation of DAG, which engaged PKCδ and IRAK1/4 to augment inflammation in 3T3-L1 adipocytes.     

Mucosal interplay among commensal and pathogenic bacteria: lessons from flagellin and Toll-like receptor 5

Toll-like receptors (TLR) detect pathogen-associated molecular patterns (PAMP) and play a crucial role in triggering immunity. Due to their large surfaces in direct contact with the environment, mucosal tissues are the major sites of PAMP-TLR signalling. How innate and adaptive immunity are triggered through flagellin-TLR5 interaction is the main focus of the review. In view of recent reports on genetic polymorphism, we will summarize the impact of TLR5 on the susceptibility to mucosal infections and on various immuno-pathologies. Finally, the contribution of TLRs in the induction and maintenance of mucosal homeostasis and commensal discrimination is discussed.     

Coordinate transformations for eye and arm movements in the brain

Recent work on the coding of spatial information in the brain has significantly advanced our knowledge of sensory to motor transformations on several fronts. The encoding of information referenced to the retina (eye-centered) but modulated by eye position, called a gain field representation, has proved to be very common throughout parietal and occipital cortex. The use of an eye-centered representation as a working memory of spatial location is problematic if the eyes move during the memory period. Details regarding the manner in which the brain solves this problem are beginning to emerge. Finally, the discovery of eye-centered representations of ongoing or intended arm movements has changed the way we think about the order of operations in the sensory to motor coordinate transformation.     

An adjuvant strategy enabled by modulation of the physical properties of microbial ligands expands antigen immunogenicity

1. Download : Download high-res image (270KB)
2. Download : Download full-size image

     

Morphological patterns of the intraparietal sulcus and the anterior intermediate parietal sulcus of Jensen in the human brain

Distinct parts of the intraparietal sulcal cortex contribute to sensorimotor integration and visual spatial attentional processing. A detailed examination of the morphological relations of the different segments of the complex intraparietal sulcal region in the human brain in standard stereotaxic space, which is a prerequisite for detailed structure-to-function studies, is not available. This study examined the intraparietal sulcus (IPS) and the related sulcus of Jensen in magnetic resonance imaging brain volumes registered in the Montreal Neurological Institute stereotaxic space. It was demonstrated that the IPS is divided into two branches: the anterior ramus and the posterior ramus of the IPS, often separated by a submerged gyral passage. The sulcus of Jensen emerges between the anterior and posterior rami of the IPS, and its ventral end is positioned between the first and second caudal branches of the superior temporal sulcus. In a small number of brains, the sulcus of Jensen may merge superficially with the first caudal branch of the superior temporal sulcus. The above morphological findings are discussed in relation to previously reported functional neuroimaging findings and provide the basis for future exploration of structure-to-function relations in the posterior parietal region of individual subjects.